Mobile Phase Of Acetonitrile Research Articles

Determination of meloxicam in human plasma by ultra-high-performance liquid chromatography-tandem mass spectrometry and its application in a pharmacokinetic study.

A rapid, selective and sensitive ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was developed to detect meloxicam in human plasma. A triple quadrupole tandem mass spectrometer equipped with an electrospray ionization source was used in positive ion mode. Protein precipitation with acetonitrile was used for sample preparation. Meloxicam and 13 C6 -meloxicam internal standard were analyzed on an Acquity CSH C18 column with a mobile phase of acetonitrile and water in 0.1% formic acid using a gradient program for separation. The retention time of meloxicam was 1.1min and the total run time was only 2.0min. Detection was performed in multiple reaction monitoring mode using an electrospray ionization source with optimized mass spectrometry parameters. The calibration curves were linear in the range 10.0-3.00 × 103 ng/ml (r ≥ 0.99). The within-run and between-run RSDs were ≤14.8%. The within-run and between-run REs ranged from -4.6 to 10.7%. There was no significant matrix effect, and the recovery rate was high. This method was fully validated, including reinjection reproducibility in human plasma. The method was applied to the pharmacokinetic study. All of the incurred sample reanalysis methods met the criteria.

Quantitative Determination of the PARP Inhibitor Olaparib (AZD 2281) in Rat Plasma using LC-MS/MS: Application to Pharmacokinetic Studies

Purpose: Olaparib, a highly selective PARP inhibitor in advanced treatment of ovarian cancer. The method describes a simple, rapid, sensitive, specific LC-ESI-MS/MS assay for the simultaneous detection and accurate measurement of olaparib in rat plasma using telmisartan as an internal standard as per the regulatory guidelines.
 Methods: Chromatographic separation was carried out on a Liquid Chromatography with tandem mass spectrometry (LC-MS/MS) unit with a Kinetex EVO C18 column (50 × 4.6 mm, 5 µ) using a gradient mobile phase of acetonitrile and 5mM ammonium acetate in water. No endogenous interfering compounds were discovered at the retention time of olaparib (1.66 min) and telmisartan (IS, 1.77 min).
 Results: The MS/MS detection was performed in positive mode and MRM transitions were m/z 435.22→366.96 and 515.21→276.16 for olaparib and IS, respectively. This method was assessed to be stable, selective and no matrix effect in three concentrations (4, 500, 800 ng/mL). The intra and inter-day precisions were less than 7.55 % and accuracy ranged from 98.00 % to 106.38 %. The extraction recovery was within acceptable limits. Additionally, the method had good linearity in the range of 1-1000 ng/mL.
 Conclusion: The validated method was successfully applied to the pharmacokinetic study of rats through oral and intravenous administration routes.

An Experimental Design Approach to Quantitative Expression for Quality Control of a Multicomponent Antidiabetic Formulation by the HILIC Method.

A rapid and reproducible hydrophilic liquid chromatography (HILIC) process was established for concomitant determination of remogliflozin etabonate (RE), vildagliptin (VD), and metformin (MF) in a formulation. A face-centered central composite experimental design was employed to optimize and predict the chromatographic condition by statistically studying the surface response model and design space with desirability close to one. A HILIC column with a simple mobile phase of acetonitrile (65% v/v) and 20 mM phosphate buffer (35% v/v, pH 6, controlled with orthophosphoric acid) was used to separate RE, VD, and MF. RE, VD, and MF were separated in 3.6 min using an isocratic mode mobile phase flow at a flow rate of 1.4 mL at room temperature, and the analytes were examined by recording the absorption at 210 nm. The developed HILIC method was thoroughly validated for all parameters recommended by ICH, and linearity was observed in the ranges 20–150 µg/mL, 10–75 µg/mL, and 50–750 µg/mL for RE, VD, and MF, respectively, along with excellent regression coefficients (r2 > 0.999). The calculated percentage relative deviation and relative error ascertained the precision and accuracy of the method. The selectivity and accuracy were further confirmed by the high percentage recovery of added standard drugs to the formulation using the standard addition technique. The robustness of the HILIC processes was confirmed by developing a half-normal probability plot and Pareto chart, as the slight variation of a single factor had no significant influence on the assay outcomes. Utilization of the optimized HILIC procedure for concurrent quantification of RE, VD, and MF in solid dosage forms showed accurate and reproducible results. Hence, the fast HILIC method can be regularly employed for the quality assurance of pharmaceutical preparations comprising RE, VD, and MF.

A Comparative Study on Analysis of Ginsenosides in American Ginseng Root Residue by HPLC-DAD-ESI-MS and UPLC-HRMS-MS/MS.

Ginseng (Panax quinquefolius), a popular herbal and nutritional supplement consumed worldwide, has been demonstrated to possess vital biological activities, which can be attributed to the presence of ginsenosides. However, the presence of ginsenosides in ginseng root residue, a by-product obtained during processing of ginseng beverage, remains unexplored. The objectives of this study were to develop a high-performance liquid chromatography-photodiode array detection-mass spectrometry (HPLC-DAD-ESI-MS) and an ultra-high-performance-liquid-chromatography-tandem mass spectrometry (UPLC-HRMS-MS/MS) method for the comparison of ginsenoside analysis in ginseng root residue. Results showed that by employing a Supelco Ascentis Express C18 column (150 × 4.6 mm ID, particle size 2.7 μm) and a gradient mobile phase of deionized water and acetonitrile with a flow rate at 1 mL/min and detection at 205 nm, a total of 10 ginsenosides, including internal standard saikosaponin A, were separated within 18 min and detected by HPLC-DAD-ESI-MS. Whereas with UPLC-HRMS-MS/MS, all the 10 ginsenosides were separated within six minutes by using an Acquity UPLC BEH C18 column (50 × 2.1 mm ID, particle size 1.7 μm, 130 Å) and a gradient mobile phase of ammonium acetate and acetonitrile with column temperature at 50 °C, flow rate at 0.4 mL/min and detection by selected reaction monitoring (SRM) mode. High accuracy and precision was shown, with limit of quantitation (LOQ) ranging from 0.2–1.9 μg/g for HPLC-DAD-ESI-MS and 0.269–6.640 ng/g for UPLC-HRMS-MS/MS. The contents of nine ginsenosides in the ginseng root residue ranged from <LOQ-26.39 mg/g by HPLC-DAD-ESI-MS and <LOQ-21.25 mg/g by UPLC-HRMS-MS/MS, with a total amount of 38.37 and 34.71 mg/g, respectively.

Pharmacokinetics and metabolism of penindolone in rat plasma using liquid chromatography–tandem mass spectrometry

Penindolone (PND) is a novel influenza A virus dual inhibitor that blocks hemagglutinin-mediated adsorption and membrane fusion. A sensitive and specific ultra-performance liquid chromatography-tandem mass spectrometry method was developed and validated to determine PND in rat plasma. Plasma sample preparation was a simple deproteinization with acetonitrile followed by centrifugation. Chromatographic separation was performed on a C18 column with a gradient mobile phase of acetonitrile-water containing 0.1% formic acid. Detection was carried out by electrospray ionization in negative ion multiple reaction monitoring mode. Linear detection responses were obtained for PND ranging from 1 to 1,000 ng/ml. The intra- and inter-day precision (relative standard deviations, RSD) were within 6.5%, and accuracy (relative error, RE) was within ±11.0%. The extraction recovery data for PND and internal standard (IS) were >96.0%. PND was proved to be stable during the sample storage, preparation and analytic procedures. The validated method was successfully applied to pharmacokinetic and bioavailability studies for PND in rats. The results showed the existence of twin peaks, gender difference and nonlinear pharmacokinetics for PND. In addition, two oxidation metabolites and three glucuronidation metabolites of PND were detected by ultra-high-performance liquid chromatography-high resolution mass spectrometry.

RP-HPLC Method development and validation for simultaneous estimation of montelukast and ebastine in pharmaceutical dosage form

Combination therapy of Montelukast (MNK) and Ebastine (EBA) provide anti-asthmatic effector the maintain treatment of asthma and relieve symptoms of allergies. The objective of this study is development of a new simple, accurate, sensitive, and reproducible RP-HPLC method for simultaneous estimation of MNK and EBA in pharmaceutical formulation (tablet) using Ofloxacin (OFL) as an internal standard and validate the same as per ICH guidelines. The chromatogram separation was achieved on Qualisil-5 BDS C8 column (250 mm × 4.6 mm, 5µm) column with mobile phase acetonitrile: water (pH 2.8 with TFA) in the composition of 84:16 v/v at a flow rate of 1 mL/min using PDA detector at 254 nm at ambient column temperature, keeping the injection volume 20 μL. The retention time of OFL, MNK, and EBA was observed to be 2.107 min, 2.517 min, and 3.819 min, respectively. All the criteria for the validation (linearity, accuracy, precision, and robustness) were observed to be within the acceptation range. The calibration plots were obtained between 5-60 µg/mL for MNK and 5-60 µg/mL for EBA with r2 values of 0.999 in each case. The recovery of MNK and EBA was found to be 98.99% and 99.40%, respectively with a % RSD of &lt;2. This RP-HPLC method was found to be rapid, specific, precise, and accurate and can be used for the routine analysis of MNK and EBA in bulk as well as in tablet dosage form. The separation was complete with a shorter analysis time along with well good resolved peak.

An in-vivo herb-drug interaction study of Tinospora cordifolia extract on the pharmacokinetics of gliclazide in normal Wistar rats

Background and Aims: Tinospora cordifolia (Willd.) Miers (TC), from the Menispermaceae family, is a well-known traditional herb used to treat diabetes mellitus and a variety of other ailments, according to traditional Indian literature. Herbal treatments are commonly used as alternatives to medicines. Anti-diabetic herbal medicines containing TC are available in the market. TC has been identified as a powerful CYP2C9 inhibitor, boosting the risk of herb-drug interactions (HDI) when used with medications metabolized via the CYP pathway. The present study evaluated the pharmacokinetic HDI of TC extract with Gliclazide (GL) after oral co-administration in Wistar rats using in vivo pharmacokinetic studies. Methods: A simple, sensitive &amp; accurate RP-HPLC/PDA method was developed and validated. Chromatographic separations were performed on a C18 analytical column with a mobile phase of acetonitrile and 40 mM ammonium acetate buffer (55:45 v/v) pH 3.0 and flow rate gradient programming. The drug concentrations in plasma were measured using the RP-HPLC method after oral co-administration of TC extract (100 mg/kg) with GL (8.3 mg/kg) in Wistar rats. Results: The pharmacokinetic (PK) interaction studies showed that the bioavailability of GL had significantly increased, with a significant influence on Cmax, AUC0-48, and suppression of volume of distribution (Vd), with no change in Tmax. Conclusion: The results obtained from this in vivo study proposed that there is a potentially significant PK HDI when GL and TC extract are administered together. This knowledge of potential HDI could be beneficial to healthcare providers and diabetic patients on GL medication. Further research is needed to anticipate this pharmacokinetics HDI of GL in humans.

Determination of Vitamin E Levels in n-Hexane Extract of Palm Fruits (Elaeis guineensis Jacq) and CPO (Crude Palm Oil) using HPLC Method

Palm fruit (Elaeis guineensis Jacq) is a natural source of vitamin E (tocopherol). This study has determined vitamin E levels in n-hexane extracts of palm fruit (Elaeis guineensis Jacq) and CPO (Crude Palm Oil) using HPLC (High Performance Liquid Chromatography) method. The subject were palm fruit and CPO obtained from plantations and palm fruit mills PT X located in Kampar district, Riau. Extraction of palm fruit and CPO using ultrasonication and saponification method. Measurement using HPLC obtained optimum conditions using mobile phase acetonitrile: water (86:14), flow 1 mL/minute, UV-Vis detector at wavelength 284 nm and obtained retention time is 3,488 minutes. Determination of the calibration curve is obtained the regression equation y = 702,872x + 3273,26 and r2 = 0,9985. The result of the determination vitamin E levels in the palm fruit extract and CPO sequentially is (469,4008 ± 2,5879) µg/mg and for CPO is (262,5248 ± 6,3987) µg/mg.

A new stability indicating RP-UFLC method for the estimation of Etoricoxib in pharmaceutical dosage forms

Etoricoxib is a selective COX-2 inhibitor and used as a non-steroidal anti-inflammatory drug. A new validated stability indicating RP-UFLC method has been developed for the estimation of Etoricoxib in pharmaceutical dosage forms (Tablets). Shimadzu UFLC system (PDA detector) with Agilent C18 column was chosen for the chromatographic study with mobile phase Acetonitrile: water (50: 50) and flow rate 0.8 mL/min (UV detection at 230 nm). Beer-Lambert’s law was obeyed over a concentration range 0.1-100 µg/mL. The linear regression equation was found to be y = 82642x -13032 (R2 = 0.9998). The LOD and LOQ was found to be and 0.0291 and 0.0894 µg/mL respectively. Stress degradation studies were performed and the method was validated as per ICH guidelines. The proposed method is simple, precise, accurate and robust and can be applied for the quantification of Etoricoxib formulations.

A Novel Method Development and Validation or the Estimation of Meloxicam in Bulk and Dosage Forms by RP – UFLC

Meloxicam is a non-steroidal anti-inflammatory drug and selective inhibitor of COX-2, it is used particularly for the management of rheumatoid arthritis. In this RP-HPLC method chromatographic separation was achieved on Isocratic elution of the mobile phase Acetonitrile: Ammonium acetate: methanol: glacial acetic acid (Proportion of mobile phase-45:40:10:5 %v/v/v/v) with the flow rate of 1 ml/min. Separation was performed on C18 column (150 mm × 4.6 mm inner diameter, 5 μm particle size). The flow rate was 1.0 ml/min and detector wavelength were kept at 363 nm for monitoring the separation. The method was developed and validated. The linearity range was found to be 10 - 50μg/ml. Regression coefficient was found to be 0.995. Precision study showed % RSD values are less than 2% in all selected concentrations. The recovery studies for Meloxicam were found to be in the range of 99.7 – 100.2%. System suitability parameters remain unchanged even if the composition of mobile phase and wavelengths were changed. The proposed method was very simple, precise, accurate and rapid for determination of Meloxicam from pure and its dosage forms.

A NEW RELATED SUBSTANCES METHOD DEVELOPMENT AND VALIDATION OF TWO ANTI-CANCER DRUGS BY USING EFFECTIVE LIQUID CHROMATOGRAPHIC METHOD

Objective: The present study was aimed at developing and successively validating novel, simple, responsive and stable RP-HPLC method for the measurement of active pharmaceutical ingredients of mitomycin and fluorouracil and their related substances. Methods: Using the impurity-spiked solution, the chromatographic approach was optimized. The chromatographic method used Luna C18 column of dimensions 150x4.6 mn, 3.5 μm, using gradient elution with a mobile phase of acetonitrile and 0.1 percent orthophosphoric acid. A flow rate of 1 ml/min and a detector wavelength of 260 nm using the PDA detector was provided in the instrumental settings. Recovery, specificity, linearity, accuracy, robustness, ruggedness was determined as a part of method validation and the results were found to be within the acceptable range. Results: According to the ICH guidelines, the developed approach was validated. The calibration charts plotted were linear with a regression coefficient of R2&gt;0.999. Conclusion: The method developed was found to be applicable to routine analysis and to be used for the measurement of active pharmaceutical ingredients (i. e, Mitomycin and Fluorouracil and its related impurities). Since there is no HPLC method reported in the literature for the estimation of Mitomycin, Fluorouracil and its related impurities; there is a need to develop quantitative methods under different conditions to achieve improvement in specificity, selectivity etc.

RAPID DETERMINATION OF CARBOPLATIN AND DOCETAXEL USİNG RP-HPLC WİTH PDA DETECTOR

Objective: In the current investigation, to separated and validate the cancer healing drugs (Carboplatin and Docetaxel) through the HPLC (e-2695) instrument containing a PDA detector. Methods: A simple, selective, validated and well-defined stability that shows isocratic RP-HPLC methodology for the quantitative determination of Carboplatin and Docetaxel. The chromatographic strategy utilized Symmetry C18 column of dimensions 150x4.6 mm, 3.5 micron, using isocratic elution with a mobile phase of acetonitrile and 0.1% ortho phosphoric acid (40:60). A flow rate of 1 ml/min and a detector wavelength of 225 nm utilizing the PDA detector were given in the instrumental settings. Recovery, specificity, linearity, accuracy, robustness, ruggedness were determined as a part of method validation and the results were found to be within the acceptable range. Validation of the proposed method was carried out according to an international conference on harmonization (ICH) guidelines. Results: LOD and LOQ for the two active ingredients were established with respect to test concentration. The calibration charts plotted were linear with a regression coefficient of R2&gt;0.999. Conclusion: The proposed method to be fast, simple, feasible and affordable in assay condition. During stability tests, it can be used for routine analysis of production samples and to verify the quality of drug samples during stability studies.

ANALYTICAL METHOD DEVELOPMENT AND VALIDATION OF DEXMETHYLPHENIDATE AND SERDEXMETHYLPHENIDATE BY USING RP-HPLC IN BULK AND PHARMACEUTICAL DOSAGE FORM

Objective: The current investigation was pointed at developing and progressively validating novel, simple, responsive and stable RP-HPLC method for the simultaneous measurement of active pharmaceutical ingredients of Dexmethylphenidate and Serdexmethylphenidate. Methods: A simple, selective, validated and well-defined stability that shows isocratic RP-HPLC methodology for the simultaneous determination of Dexmethylphenidate and Serdexmethylphenidate. The chromatographic strategy utilized inertsil ODS column of dimensions 250x4.6 mm, 5 µ, using isocratic elution with a mobile phase of acetonitrile and 0.1% orthophosphoric acid (70:30). A flow rate of 1 ml/min and a detector wavelength of 262 nm utilizing the PDA detector were given in the instrumental settings. Recovery, specificity, linearity, accuracy, robustness, ruggedness were determined as a part of method validation and the results were found to be within the acceptable range. Validation of the proposed method was carried out according to an international conference on harmonization (ICH) guidelines. Results: LOD and LOQ for the active ingredient were established with respect to test concentration. The calibration chart plotted was linear with a regression coefficient of R2&gt;0.999, which means the linearity was within the limit. Conclusion: The proposed method to be fast, simple, feasible and affordable in assay condition. During stability tests, it can be used for routine analysis of production samples and to verify the quality of drug samples during stability studies.

ASSAY METHOD DEVELOPMENT AND VALIDATION OF CILNIDIPINE AND RAMIPRIL, CHARACTERIZATION OF ITS DEGRADANTS BY USING LC-MS/MS

Objective: The current study focused on the development, validation of Ramipril and Cilnidipine using HPLC and characterization of forced degradation products using LC-MS/MS. Methods: Simple, accurate, and precise method has been developed for the simultaneous estimation of Cilnidipine and Ramipril in tablet type of dosage form using X-bridge phenyl column (150x4.6 mm, 3.5m) with mobile phase of buffer and Acetonitrile in the 30:70 v/v ratio was pumped through a column with a 1 ml/min flow rate. The buffer used in this process was 0.1 percent tri ethyl amine in 1 lt of water and adjust its pH-2.5 with 0.1 percent ortho phosphoric acid. At ambient temperature, chromatography was isocratically performed and run time was 8 min. 240 nm was the optimized wave length. Results: Cilnidipine and Ramipril retention times were 2.79 min, 5.11 min. respectively. By injecting the norm six times, device parameters of suitability have been studied and the results were found to be well under the acceptance criteria. This approach offers strong linearity over a range of 2-30 µg/ml Cilnidipine 1-15 µg/ml Ramipril concentrations. The regression coefficient R2&gt;0.999 from the calibrated curve implies that the linearity of the system was within the range. Conclusion: The system was validated by using HPLC in terms of accuracy, linearity, method precision, accuracy, limit of detection, limit of quantification, robustness, degradation and the degradation products were characterized by using LC-MS/MS.

Cucurbit[8]uril-Based Potentiometric Sensor Coupled to HPLC for Determination of Tetracycline Residues in Milk Samples

The determination of chlortetracycline, doxycycline, oxytetracycline, and tetracycline in milk samples by HPLC coupled to a cucurbit[8]uril-based potentiometric sensor is herein presented. The new tetracycline-selective electrode is based on a polymeric membrane incorporating cucurbit[8]uril as a macrocyclic host, potassium tetrakis(p-chlorophenyl) borate as an ionic additive, 2-fluorophenyl 2-nitrophenyl ether as a plasticizer, and multi-walled carbon nanotubes as nanostructured materials. A microfluidic wall-jet flow-cell is implemented as a potentiometric detector after chromatographic separation by a C8 column using a gradient mobile phase of sulphuric acid and acetonitrile. The proposed methodology was validated following International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) and European Union (EU) guidelines. Linear regression models provided R2 in the range from 0.9973 ± 0.0026 to 0.9987 ± 0.0012 for all tetracycline antibiotics. The limits of detection and quantification ranged from 13.3 to 46.0 μg L−1 and 44.4 to 92.1 μg L−1, respectively. Precision intra-day, inter-day, and inter-electrode showed relative standard deviation values lower than 12.5%, 13.5%, and 12.9%, respectively. Accuracy was assessed by analysis of spiked milk samples around the maximum residue limit, yielding recovery values in the range from 81.3 to 108.5%. The simple, sensitive, cost-effective, and reliable HPLC-ion-selective electrode method justifies its use as a competitive alternative for the analysis of tetracycline residues in the food quality control sector.

Retention properties of acetone-water mobile phases on a biphenylsiloxane-bonded silica stationary phase in reversed-phase liquid chromatography.

The solvation parameter model system constants and retention factors were used to interpret retention properties of 39 calibration compounds on a biphenylsiloxane-bonded stationary phase (Kinetex biphenyl) for acetone-water binary mobile phase systems containing 30-70% v/v. Variation in system constants, phase ratios, and retention factors of acetone-water binary mobile phases systems were compared with more commonly used acetonitrile and methanol mobile phase systems. Retention properties of acetone mobile phases on a Kinetex biphenyl column were more similar to that of acetonitrile than methanol mobile phases except with respect to selectivity equivalency. Importantly, selectivity differences arising between acetone and acetonitrile systems (the lower hydrogen-bond basicity of acetone-water mobile phases and differences in hydrogen-bond acidity, cavity formation and dispersion interactions) could be exploited in reversed-phase liquid chromatography method development on a Kinetex biphenyl stationary phase.

Development and validation of RP-HPLC method for analytical characterization of the anabolic steroid Methenolone acetate in food supplements

Nutritional supplements are concentrated sources of nutrients, vitamins and minerals with a nutritional or physiological effect, the purpose of which is to supplement the normal nutritional balance. A necessity of the health risk associated with their unregulated usage requires development of suitable fast and precise methods for their evaluation. As an answer to this a novel RP-HPLC method for analytical characterization of the anabolic steroid Methenolone acetate in food supplement was developed. The method is based on separation and evaluation performed on an analytical column Kintex 5µm EVO C18 (100 × 4.6 mm) with mobile phase acetonitrile:water = 60:40, v/v; flow rate of 1.0 mL/min and UV-detection at 240 nm and column temperature, at 25 °C. The method has proved to be specific, linear, accurate and precise. The method identified a presence of both Methenolone and Methenolone acetate in the evaluated sample.

Simultaneous Analysis for Quality Control of Traditional Herbal Medicine, Gungha-Tang, Using Liquid Chromatography-Tandem Mass Spectrometry.

Gungha-tang (GHT), a traditional herbal medicine, consists of nine medicinal herbs (Cnidii Rhizoma, Pinelliae Tuber, Poria Sclerotium, Citri Unshius Pericarpium, Citri Unshius Pericarpium Immaturus, Aurantii Fructus Immaturus, Atracylodis Rhizoma Alba, Glycyrrhizae Radix et Rhizoma, and Zingiberis Rhizoma Recens). It has been used for various diseases caused by phlegm. This study aimed to develop and verify the simultaneous liquid chromatography–tandem mass spectrometry (LC–MS/MS) analysis method, using nine marker components (liquiritin apioside, neoeriocitrin, narirutin, naringin, hesperidin, neohesperidin, liquiritigenin, glycyrrhizin, and 6-shogaol) for quality control of GHT. LC–MS/MS analysis was conducted using a Waters TQ-XS system. All marker analytes were separated on a Waters Acquity UPLC BEH C18 column (2.1 × 100 mm, 1.7 μm) using gradient elution with a distilled water solution (containing 5 mM ammonium formate and 0.1% [v/v] formic acid)–acetonitrile mobile phase. LC–MS/MS multiple reaction monitoring (MRM) analysis was carried out in negative and positive ion modes of an electrospray ionization source. The developed LC–MS/MS MRM method was validated by examining the linearity, limits of detection and quantification, recovery, and precision. LOD and LOQ values of nine markers were calculated as 0.02–8.33 ng/mL and 0.05–25.00 ng/mL. The recovery was determined to be 89.00–118.08% and precision was assessed with a coefficient of variation value of 1.74–8.64%. In the established LC–MS/MS MRM method, all markers in GHT samples were detected at 0.003–16.157 mg/g. Information gathered during the development and verification of the LC–MS/MS method will be useful for the quality assessment of GHT and other herbal medicines.

RP-HPLC-UV validation method for levofloxacin hemihydrate estimation in the nano polymeric ocular preparation

The specific and accurate reversed-phase HPLC-UV method has been validated to determine levofloxacin hemihydrate (LEVH) level. The separation was conducted at C 18 analytical column by administering mobile phase acetonitrile, methanol, and phosphate buffer (pH 3) with the ratio of 17:3:80. The flow rate of the mobile phase was 1 mL/min with a UV detector at 295 nm wavelength. Analytical methods validation evaluated includes specificity, linearity, accuracy, precision, LOD, LOQ, and robustness. The implementation of the analytical method was employed to determine LEVH level in ocular polymeric nanoparticles preparations. The test was specific for LEVH with the retention time of 7.66 min. Linearity was obtained from the concentration range of 4.8 µg/mL to 29.04 µg/mL. All method validation criteria are within the acceptable range. The developed method can be applied for LEVH polymeric nano-formulation analysis.

Development and validation of the high-performance liquid chromatography method for estimating N-(n-butyl) thiophosphoric triamide in granular fertilisers

N-(n-butyl) thiophosphoric triamide is widely used as a urease inhibitor that can reduce nitrogen loss by ammonia evaporation from urea. It is used as a fertiliser additive in agricultural applications. The recommended doses of inhibitors are not dangerous to animals and the environment, but high doses can cause changes in small animals. Therefore, the amount of inhibitors in the fertiliser must be determined before fertilising the soil. This study developed and validated a reliable, sensitive, rapid, and precise high-performance liquid chromatography method for the quantification of N-(n-butyl) thiophosphoric triamide in granular fertilisers. The estimation was carried out using a YMC-Triat C18 column (150 × 4.6 mm, 3 mm) and the mobile phase of acetonitrile: deionised water (25:75, v/v). Inhibition efficiency was monitored by a UV detector at 205 nm. The total run time was 10 min with a flow rate of 0.8 mL/min. The parameters considered for validation were linearity, detection and quantification limit, sustainability, robustness, precision, and stability. The newly created innovative method for quantitative estimation of N-(n-butyl) thiophosphoric triamide in granular fertilisers improves the solubility of samples and standard substance the repeatability and reproducibility of the results obtained.